Defining the function of Schlafen4 in the pathogenesis of flavivirus encephalitis Abstract: Members of flavivirus genus are the most important arthropod-borne viruses causing disease in humans. This genus includes West Nile virus (WNV) and Japanese encephalitis virus (JEV) that are the leading causes of arboviral encephalitis in the United States and worldwide. No effective therapies exist for treating individuals with encephalitic flavivirus infections, and pathogenesis of flavivirus encephalitis is not completely understood. Addressing the fundamental questions regarding host proteins recruited by the flaviviruses to establish infection in the primary target cells is an integrated and indispensable part of the effort towards development of effective therapeutics. Schlafen4 (SLFN4) is a poorly characterized but important member of the Schlafen family that includes several mouse and human genes. Recently we demonstrated that SLFN4 is required for WNV replication in primary mouse embryonic fibroblasts. The principal objective of the proposed exploratory research is to define the function of SLFN4 in WNV and JEV replication and pathogenesis, using novel mouse models. Given that this is the first study of its kind, aim 1 will use wild-type (WT) and SLFN4 knockout mice to investigate the function of SLFN4 in WNV and JEV replication, and disease outcome. Aim 2 will investigate the cell-specific function of SLFN4 in WNV and JEV replication using a set of conditional knockout mice with SLFN4 deleted in a cell type restricted manner in dendritic cells and macrophages, as well as using mouse cells of myeloid lineage and brain cells from WT and SLFN4-/- mice. Additionally, we will generate a knockdown of SLFN4 orthologs (SLFN12 and SLFN12L) in human monocytes, dendritic cells and neurons, and examine the kinetics of WNV and JEV replication. The proposed research is highly innovative in its use of the newly developed constitutive and cell-specific SLFN4 knockout mice models to investigate the function of SLFN4 in the pathogenesis of flavivirus encephalitis. This study will be the first study to define the functional role of this uncharacterized host factor, SLFN4, in viral infection and disease pathogenesis. Overall, our studies will provide new insight into a novel host factor for flavivirus replication and dissemination, and thus will have a significant impact on the development of much-needed therapeutic interventions that will reduce virus spread and improve disease outcome.